RNA Structural Determinants of Optimal Codons Revealed by MAGE-Seq.
RNA Structural Determinants of Optimal Codons Revealed by MAGE-Seq.
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DOI：
10.1016/j.cels.2016.11.004
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发表时间：
2016-12-21
期刊：
影响因子：
9.3
通讯作者：
Kishony R
中科院分区：
文献类型：
--
作者：
Kelsic ED;Chung H;Cohen N;Park J;Wang HH;Kishony R
基因开头的同义密码子选择优化了5'RNA结构，以增强翻译起始，但对驱动下游密码子优化的机制少知道，以了解基因跨基因的密码子选择的原因，我们在原位密码子突变体中生成了12,726在大肠杆菌本质基因Infa中，通过将多重自动基因组工程诱变与Amplicon结合在一起，测量了其适应性测序（Mage-seq）。限制固有的密码子偏好超出基因组RNA折叠预测的选择在5'UTR附近的天然结构的破坏。
Kelsic等人开发并应用法师序列来识别确定基本大肠杆菌基因中最佳密码子偏好的RNA结构。
Synonymous codon choices at the beginning of genes optimize 5′ RNA structures for enhanced translation initiation, but less is known about mechanisms that drive codon optimization downstream within the gene. To understand what determines codon choices across a gene, we generated 12,726 in situ codon mutants in the Escherichia coli essential gene infA and measured their fitness by combining multiplex automated genome engineering mutagenesis with amplicon deep sequencing (MAGE-seq). Correlating predicted 5′ RNA structure with fitness revealed that codons even far from the start of the gene are deleterious if they disrupt the native 5′ RNA conformation. These long-range structural interactions generate context-dependent rules that constrain codon choices beyond intrinsic codon preferences. Genome-wide RNA folding predictions confirm that natural codon choices far from the start codon are optimized in part to prevent disruption of native structures near the 5′ UTR. Our results shed light on natural codon distributions and should improve engineering of gene expression for synthetic biology applications. Kelsic et al. develop and apply MAGE-seq to identify RNA structures that determine optimal codon preferences in an essential E. coli gene.